KR101849119B1 - Antioxidant composition comprising fermented Bamboo Leaves - Google Patents

Abstract

The present invention relates to a composition having antioxidative activities. More specifically, the present invention relates to a functional health food and an antioxidant composition for drugs, foods, or cosmetic products, containing a fermented bamboo leaves as an active ingredient. The fermented bamboo leaves ensure enhanced content of phenolic compounds and flavonoid compounds by fermenting Sasa coreana Nakai leaves using a Bacillus subtilis strain as a specific microorganism. Since the fermented bamboo leaves exhibit excellent antioxidative activities due to an increase in phytochemicals, the composition containing the fermented bamboo leaves as the active ingredient can be useful as an antioxidant in fields of drugs, health functional foods and cosmetic products.

Description

An antioxidant composition comprising fermented bamboo leaves as an active ingredient (Fermented Bamboo Leaves)

The present invention relates to a composition having an antioxidative activity, and more particularly, to an antioxidative composition and a health functional food for pharmaceutical use, food use or cosmetic use containing a bamboo leaf fermented product as an effective ingredient.

There are 10 species of bamboo in Korea, which grows well in the southern part of the country, and is composed of Phyllostachys and Sasa species. Bamboo is a highly utilized forest resource throughout the real life, and is also used as a building material. It has a wide variety of applications such as furniture and bamboo crafts. Among the various parts of bamboo, bamboo leaves are used not only in Korea but also in processed foods such as Jatropha, Meat Leaf, and Kanebo Salt in Asian regions such as China and Japan.

Bamboo leaves contain carbohydrates, minerals, and amino acids as nutrients. They contain a variety of nutrients such as orientin, homoorientin, vitexin, isovitexin, and linoleic acid. Phytochemicals are included and exhibit various physiological activities. Recently, various attempts have been made to utilize bamboo leaves rich in such nutrients and phytochemicals as food materials and utilize the extracts. However, bamboo leaves are often extracted by taking tea or drink form rather than taking bamboo leaves themselves because of their high cellulose content, low satiety, and unique flavor.

On the other hand, fermentation is one of the processing methods that can affect the taste and flavor of food and enhance the storage stability. It is decomposed into a form in which the nutrients of the food are easily digested by the enzymatic action derived from the fermenting microorganism, You can give. Traditional fermented food manufacturing methods are difficult to obtain uniform quality because microorganisms are not identical due to regions, manufacturing methods, raw materials, etc. In order to produce high-quality fermented foods, it is important to select and use high-performance microorganisms.

In contrast to the study on nutrients of the bamboo leaves containing various physiologically active substances, studies on the changes of the components before and after the fermentation and the changes of the physiological activities when the fermentation process was applied are insufficient. In particular, There have been no studies on the changes of components and antioxidant activity by fermentation with specific microorganisms.

Accordingly, the present inventors have found that a bamboo leaf containing various physiologically active substances can be used as a naturally occurring bamboo leaf, coreana The contents of phenolic compounds and flavonoids were analyzed and their antioxidant activities were analyzed. The contents of antioxidant activity of the Nakai leaves were investigated by fermentation using Bacillus subtilis as a specific microorganism. As a result, the present inventors have completed the present invention by confirming that the antioxidative activity of the ginseng fermented with Bacillus subtilis is remarkably increased in the fermented product of the leaves.

Korean Patent Publication No. 10-2014-0086434

Accordingly, an object of the present invention is to provide an antioxidant composition for pharmaceutical use, food use, cosmetic use, and animal feed use having excellent antioxidant activity.

It is still another object of the present invention to provide a health functional food having excellent antioxidant activity.

In order to achieve the above-mentioned object of the present invention,

The present invention provides an antioxidative composition comprising a bamboo leaf fermented product as an active ingredient.

In one embodiment of the present invention, the bamboo may be Sasa coreana Nakai .

In one embodiment of the present invention, the bamboo leaf fermented product can be produced by inoculating a bamboo leaf with a Bacillus subtilis strain to ferment.

In one embodiment of the present invention, the composition may be used for medicines, foods, cosmetics, or feedstuffs.

In addition, the present invention provides a health functional food comprising the bamboo leaf fermented product as an active ingredient.

The bamboo leaf fermented product of the present invention has an increased antioxidative activity through the increase of the phytochemical by increasing the content of the phenolic compound and the flavonoid compound by fermenting the bovine leaf with the Bacillus subtilis strain as a specific microorganism, The composition comprising the bamboo leaf fermented product as an active ingredient can be usefully used as an antioxidant in the fields of pharmaceuticals, health functional foods and cosmetics.

FIG. 1 is a photograph showing the color change of the larval leaf during fermentation using Bacillus subtilis strain.
FIG. 2 is a graph showing the content of total polyphenols, total flavonoids and catechins in the extracts before fermentation (SC-N) and after (SC-F)
FIG. 3 is a graph showing antioxidative activities of SC-N and SC-F extracts before and after fermentation of Shinyi leaves.

The present invention is characterized by providing an antioxidant composition comprising a bamboo leaf fermented product as an active ingredient.

The 'bamboo leaf fermented product' referred to in the present specification is a substance obtained by fermenting bamboo leaves by inoculating fermented microorganisms on bamboo leaves. The fermented material itself of bamboo leaves, bamboo leaf fermented extract extracted with a solvent thereto, and other tablets And all extracts, fractions, tablets and the like that can be obtained through the processing process.

The 'extract' defined in the present invention is extracted from the bamboo leaf fermentation product using an appropriate solvent and includes, for example, a crude extract, a polar solvent extraction extract or a non-polar solvent extraction extract of the bamboo leaf fermentation product.

As an appropriate solvent for extracting the extract from the bamboo leaf fermented product, any pharmaceutically acceptable organic solvent may be used, and water or an organic solvent may be used. Examples of the solvent include, but are not limited to, purified water Alcohol having 1 to 4 carbon atoms, acetone, ether, benzene, and the like, including methanol, ethanol, propanol, isopropanol, butanol, Various solvents such as chloroform, ethyl acetate, methylene chloride, hexane, and cyclohexane may be used alone or in combination. It is preferable to use ethanol (alcohol), more preferably ethanol.

As the extraction method, any one of the methods such as hot water extraction method, cold extraction method, reflux cooling extraction method, solvent extraction method, steam distillation method, ultrasonic extraction method, elution method and compression method can be selected and used. In addition, the desired extract may be further subjected to a conventional fractionation process or may be purified using a conventional purification method. The method for producing the extract of the present invention is not limited, and any known method can be used.

For example, the bamboo leaf fermentation extract contained in the composition of the present invention may be prepared by subjecting a fermented bamboo leaf to an additional process such as vacuum distillation, freeze drying, or spray drying, In a powder state. In addition, the primary extract can be further fractionated using a variety of chromatographies, such as silica gel column chromatography, thin layer chromatography, high performance liquid chromatography, .

Therefore, the term " bamboo leaf fermented product " in the present invention refers to any fermented product of bamboo leaves, as well as any extract, fraction and purified product obtained by extracting, fractionating or refining the fermented product, a diluted solution thereof, It is a concept that includes everything.

In one embodiment of the present invention, the bamboo leaf may be selected from the group consisting of Sasa coreana Nakai leaves can be used.

As used herein, " Shin- iida " coreana It is a Korean bamboo that grows at the foot of the mountains such as the Nakai , the pizza plant gate, the terminal plant river, the flower head, the rice paddy field, The basement is divided at the end. Stems live for about four years, usually two years later, they branch at the top. The first-year stem is 25-80cm high, the length between the nodes is 30-70mm, the diameter is 3mm, and there are round and round hairs. The node is relatively flat and has a cuprous hairs. The length of the cauliflower is 35 ~ 75mm and it remains for a long time. There are dark hairs between the veins, but there are no hairs on the above. The leaves are 5 ~ 8 in 2 lines at the end of the stem and have 2 ~ 7mm petiole (petiole). Leaf is ovate long elliptical or elliptical, 25 ~ 190mm long, 8 ~ 50mm wide, pointed long, hairless and green on the front, Silk hair) is covered, and there is a piercing streak on the outskirts. On both sides of the leaf plot there are 6 ~ 10 veins (veins) in the vertical direction. The leaf tongue is 0.8 ~ 2mm long and has many hairs. Both the nodules and the nodules have hairs that are glowing. Leaves from a biennial branch are 180 cm long and less than 30 cm wide and have a sharp pointed base with only fine hairs on the mid ribs of the back. The leaves of 3-year-old branches are 125mm long and 30mm wide. There are no hairs on the back side, or there is a little honey hair on the bottom of the midrib, but they disappear immediately. There is no powder on the nodes. The leaves of 4-year-old branches are usually 57 ~ 120mm long and 12 ~ 20mm wide, but rarely smaller than others. They have long pointed lower part and bristles on the outskirts. Ornamental use, leaves are written in medicine.

In another embodiment of the present invention, the bamboo leaf fermented product can be produced by inoculating a bamboo leaf with a Bacillus subtilis strain to ferment.

Bacillus subtilis is a type of bacterium belonging to the genus Bacillus, and is a gram-positive bacterium widely distributed in the natural world such as soil, hay, and dust, and has a length of 2 to 3 μm, It is also called Bacillus. Spores are resistant to heat, radiation and chemicals, and remain dormant for long periods. Since the transgenic plants were able to grow in a simple medium, they were widely used for molecular genetics and recombinant DNA experiments. Bacillus subtilis is an important microorganism for producing microorganisms such as amylase and protease of extracellular enzymes, and has recently attracted attention as a material for gene engineering.

As used herein, the term "antioxidant composition" means a substance that prevents oxidation through the action of inhibiting or eliminating active oxygen in an unstable state of oxygen.

The antioxidant composition of the present invention can be used for various purposes and applications requiring an antioxidative activity. Specifically, the antioxidant composition of the present invention can be used as a functional material capable of imparting antioxidative activity to articles applied in various industrial fields such as medicines, cosmetics, And can also be used as a medicine preservative, a cosmetic preservative, a food preservative, a pharmaceutical additive, a cosmetic additive, a food additive, and a feed additive.

Hereinafter, the antioxidant composition of the present invention can be applied to pharmaceuticals, cosmetics, foods, and animal feeds in detail.

The antioxidant composition of the present invention may be a pharmaceutical composition for preventing or treating an oxidation-related disease, which comprises a bamboo leaf fermented product as an active ingredient.

The pharmaceutical composition of the present invention can be prepared by using pharmaceutically acceptable and physiologically acceptable adjuvants in addition to the above-mentioned active ingredients. Examples of the adjuvants include excipients, disintegrants, sweeteners, binders, coating agents, swelling agents, lubricants, A lubricant or a flavoring agent can be used.

The pharmaceutical composition may be formulated into a pharmaceutical composition containing at least one pharmaceutically acceptable carrier in addition to the above-described active ingredients for administration.

The pharmaceutical composition may be in the form of granules, powders, tablets, coated tablets, capsules, suppositories, liquids, syrups, juices, suspensions, emulsions, drops or injectable solutions. For example, for formulation into tablets or capsules, the active ingredient may be combined with an oral, non-toxic pharmaceutically acceptable inert carrier such as ethanol, glycerol, water, and the like. Also, if desired or necessary, suitable binders, lubricants, disintegrants and coloring agents may also be included as a mixture. Suitable binders include, but are not limited to, natural sugars such as starch, gelatin, glucose or beta-lactose, natural and synthetic gums such as corn sweeteners, acacia, tracker candles or sodium oleate, sodium stearate, magnesium stearate, sodium Benzoate, sodium acetate, sodium chloride, and the like. Disintegrants include, but are not limited to, starch, methyl cellulose, agar, bentonite, xanthan gum and the like. Acceptable pharmaceutical carriers for compositions that are formulated into a liquid solution include sterile solutions suitable for the living body such as saline, sterile water, Ringer's solution, buffered saline, albumin injection solution, dextrose solution, maltodextrin solution, glycerol, One or more of these components may be mixed and used. If necessary, other conventional additives such as an antioxidant, a buffer, and a bacteriostatic agent may be added. In addition, diluents, dispersants, surfactants, binders, and lubricants may be additionally added to formulate into injectable solutions, pills, capsules, granules or tablets such as aqueous solutions, suspensions, emulsions and the like. Further, it can be suitably formulated according to each disease or ingredient, using the method disclosed in Remington's Pharmaceutical Science, Mack Publishing Company, Easton PA as an appropriate method in the field.

In one embodiment of the present invention, the bamboo leaf fermented product of the present invention may be contained in an amount of 0.00001 to 40% by weight based on the total weight of the composition.

In another embodiment of the present invention, the bamboo leaf fermented product of the present invention may be contained at a concentration of 0.1 to 1000 μg / ml with respect to the composition.

Examples of the oxidation-related diseases in which the pharmaceutical composition of the present invention can exhibit therapeutic effects include rhinitis, asthma, acute pain, chronic pain, periodontitis, gingivitis, inflammatory bowel disease, gout, myocardial infarction, arteriosclerosis, congestive heart failure, hypertension , Angina pectoris, stomach ulcer, Alzheimer's disease, cerebral infarction, Down's syndrome, multiple sclerosis, obesity, diabetes, dementia, depression, schizophrenia, tuberculosis, sleep disorders, sepsis, burns, pancreatitis or ophthalmic diseases no.

The composition of the present invention may also be a food composition. In addition to containing the active ingredient bamboo leaf fermented product, such a food composition may contain various flavors or natural carbohydrates as an additional component such as a conventional food composition .

Examples of the above-mentioned natural carbohydrates include monosaccharides such as glucose, fructose and the like; Disaccharides such as maltose, sucrose and the like; And polysaccharides, for example, conventional sugars such as dextrin, cyclodextrin and the like, and sugar alcohols such as xylitol, sorbitol and erythritol. The above-described flavors can be advantageously used as natural flavorings (tau martin), stevia extracts (e.g., rebaudioside A, glycyrrhizin, etc.) and synthetic flavors (saccharin, aspartame, etc.).

The food composition of the present invention can be formulated in the same manner as the above pharmaceutical composition and used as a functional food or added to various foods. Foods to which the composition of the present invention can be added include, for example, beverages, meat, chocolates, foods, confectionery, pizza, ram noodles, other noodles, gums, candy, ice cream, alcoholic beverages, vitamin complexes, .

In addition to the bamboo leaf fermentation product as an active ingredient, the above food composition may also contain various nutrients, vitamins, minerals (electrolytes), flavors such as synthetic flavors and natural flavors, colorants and heavies such as cheese and chocolate, And carbonates used in alginic acid and its salts, alginic acid and its salts, organic acids, protective colloid thickeners, pH adjusting agents, stabilizers, preservatives, glycerin, alcohols, carbonated drinks and the like. In addition, the food composition of the present invention may contain natural fruit juice and pulp for the production of fruit juice drinks and vegetable drinks.

Since the bamboo leaf fermented product of the present invention is a natural substance and has almost no side effects such as chemical agents, it can be safely used for long-term administration for the purpose of imparting antioxidant function.

The present invention may also be a health functional food for preventing or ameliorating an oxidation-related disease comprising a bamboo leaf fermented product as an active ingredient.

The health functional food of the present invention can be manufactured and processed in the form of tablets, capsules, powders, granules, liquids, and rings for the purpose of preventing or improving an oxidation-related disease.

In the present invention, the term " health functional food " refers to a food prepared or processed using raw materials or ingredients having useful functions in accordance with the Act on Health Functional Foods, and the nutrients are controlled Physiological action, etc., for the purpose of obtaining a beneficial effect.

The health functional foods of the present invention may contain conventional food additives and, unless otherwise specified, whether or not they are suitable as food additives are classified according to the General Rules for Food Additives approved by the Food and Drug Administration, Standards and standards.

Examples of the items listed in the above-mentioned "food additives" include chemical compounds such as ketones, glycine, calcium citrate, nicotinic acid, and cinnamic acid; Natural additives such as persimmon extract, licorice extract, crystalline cellulose, high color pigment and guar gum; L-glutamic acid sodium preparations, noodle-added alkalis, preservative preparations, tar coloring preparations and the like.

For example, the health functional food in tablet form can be obtained by granulating a mixture of bamboo leaf fermented product as an active ingredient of the present invention with an excipient, a binder, a disintegrant and other additives in a usual manner, Compression molding, or direct compression molding of the mixture. In addition, the health functional food of the tablet form may contain a mating agent or the like if necessary.

The hard capsule of the capsule type health functional food can be prepared by filling a normal hard capsule with a mixture of the bamboo leaf fermented product as an active ingredient of the present invention with an additive such as an excipient and the soft capsule may contain the active ingredient The bamboo leaf fermented product is mixed with an additive such as an excipient and the mixture is filled in a capsule base such as gelatin. The soft capsule may contain a plasticizer such as glycerin or sorbitol, a coloring agent, a preservative and the like, if necessary.

The ring-shaped health functional food can be prepared by molding a mixture of the bamboo leaf fermented product of the present invention, excipient, binder, disintegrant and the like by a conventionally known method, and if necessary, The surface can be coated with a material such as starch, talc or the like.

The granular health functional food may be prepared by granulating a mixture of the bamboo leaf fermented product of the present invention, excipient, binder, disintegrant, and the like, into granules by a known method, and if necessary, And the like.

The health functional food containing the bamboo leaf fermented product of the present invention as an active ingredient has proven its excellent antioxidative effect as shown in the following examples, and thus it is effective for prevention or improvement of oxidation-related diseases.

The health functional food may be a beverage, a meat, a chocolate, a food, a confectionery, a pizza, a ramen, a noodle, a gum, a candy, an ice cream, an alcoholic beverage, a vitamin complex and a health supplement food.

In addition, the composition of the present invention may be a cosmetic composition in addition to a pharmaceutical composition or a food composition.

The bamboo leaf fermented product of the present invention has excellent antioxidant activity because it has excellent scavenging ability against hydroxyl radicals having the highest reactivity among active oxygen species as well as nitric oxide scavenging activity. Therefore, a composition comprising the bamboo leaf fermented product as an active ingredient Can be effectively used as a cosmetic composition because it is effective for skin protection and skin treatment from active oxygen species (ROS) that promote damage to human cells and accelerate aging and mediate oxidation-related diseases.

When the composition of the present invention is prepared with a cosmetic composition, the composition of the present invention may contain not only the above-mentioned bamboo leaf fermented product, but also components commonly used in cosmetic compositions such as antioxidants, stabilizers, Vitamins, pigments and flavoring agents, and carriers.

In addition to the bamboo leaf fermentation product of the present invention, the composition of the present invention may be mixed with an antioxidant which has been used in the past so long as it does not impair the skin protecting effect by reacting with the bamboo leaf fermentation product.

Examples of products to which the cosmetic composition of the present invention can be added include cosmetics such as astringent lotion, softening longevity lotion, nutrition lotion, various creams, essences, packs, foundation and the like, cleansing, cleanser, soap, .

As a specific formulation of the cosmetic composition of the present invention, there may be mentioned a skin lotion, a skin softener, a skin toner, an astringent, a lotion, a milk lotion, a moisturizing lotion, a nutrition lotion, a massage cream, It includes formulations such as soap, shampoo, cleansing foam, cleansing lotion, cleansing cream, body lotion, body cleanser, latex, lipstick, makeup base, foundation, press powder, loose powder, eye shadow.

According to a preferred embodiment of the present invention, the content of the bamboo leaf fermentation product of the present invention is 0.00001-40 wt%, preferably 0.1-20 wt%, more preferably 1.0-20 wt%, based on the total weight of the composition. If the content of the bamboo leaf fermented product is less than 0.00001% by weight, the antioxidative effect of the extract and its fractions is greatly reduced. If the content is more than 40% by weight, skin irritation may be caused.

Meanwhile, the cosmetic composition according to the present invention may be formulated by stabilizing the bamboo leaf fermented product by containing it in the nanoliposome. When the extract (or fraction thereof) is contained in the inside of the nanoliposome, the components of the extract (or the fraction thereof) are stabilized to solve problems such as precipitation, discoloration, and deterioration upon formulation, and the solubility and transdermal absorption To maximize the efficacy expected from the extract (or fractions thereof).

In the present invention, nanoliposome refers to a liposome having a typical liposome form and having an average particle diameter of 10 to 500 nm. According to a preferred embodiment of the present invention, the average particle diameter of the nanoliposome is 50 to 300 nm. When the average particle diameter of the nanoliposome is more than 300 nm, improvement of skin penetration and improvement of formulation stability is very weak among technological effects to be achieved in the present invention.

The nanoliposomes used to stabilize the extract (or fractions thereof) according to the present invention may be prepared by a mixture comprising a polyol, an oily component, a surfactant, a phospholipid, a fatty acid and water.

The polyol used in the nanoliposome of the present invention is not particularly limited and is preferably a polyol such as propylene glycol, dipropylene glycol, 1,3-butylene glycol, glycerin, methylpropanediol, isopropylene glycol, pentylene glycol, erythritol, , Sorbitol, and mixtures thereof. The amount thereof is 10 to 80% by weight, preferably 30 to 70% by weight, based on the total weight of the nanoliposome.

The oil component used in the preparation of the nanoliposome of the present invention may be selected from a variety of oils known in the art and is preferably a hydrocarbon oil such as hexadecane and paraffin oil, Vegetable oils such as silicon oil such as dimethicone and cyclomethicone, sunflower oil, corn oil, soybean oil, avocado oil, sesame oil and fish oil, ethoxylated alkyl ether oils, propoxylated alkyl ether oils ₄₀ is a fatty alcohol, and mixtures thereof -, phytosphingosine, sphingosine, and scan non-ping the same scan pinggo cannabinoid lipid, cholesterol side-by celebrity, sitosterol cholesteryl sulfate, cholesteryl sulfate, cytokines, C _10. The amount thereof may be 1.0 to 30.0% by weight, and preferably 3.0 to 20.0% by weight based on the total weight of the nanoliposome.

Any surfactant known in the art may be used as the surfactant used in the preparation of the nanoliposome of the present invention. For example, anionic surfactants, cationic surfactants, amphoteric surfactants and nonionic surfactants can be used. Preferred are anionic surfactants and nonionic surfactants. Specific examples of anionic surfactants include alkyl acyl glutamates, alkyl phosphates, alkyl lactylates, dialkyl phosphates and trialkyl phosphates. Specific examples of nonionic surfactants include alkoxylated alkyl ethers, alkoxylated alkyl esters, alkyl polyglycosides, polyglyceryl esters and sugar esters. Particularly preferred surfactants are polysorbates belonging to nonionic surfactants. The amount thereof may be 0.1 to 10% by weight, preferably 0.5 to 5.0% by weight, based on the total weight of the nanoliposome.

The phospholipid, another component used in the preparation of the nanoliposomes of the present invention, is a lipophilic lipid that is used with both affinity lipids and is composed of natural phospholipids (e.g., egg yolk lecithin or soy lecithin, sphingomyelin) Dipalmitoyl phosphatidylcholine or hydrogenated lecithin), preferably lecithin. In particular, unsaturated lecithin or saturated lecithin derived from natural origin extracted from soybean or egg yolk is preferable. Normally, the amount of phosphatidylcholine is 23 to 95% and the amount of phosphatidylethanolamine is 20% or less in natural derived lecithin. In the production of the nanoliposome of the present invention, the amount of the phospholipid to be used is 0.5 to 20.0% by weight, preferably 2.0 to 8.0% by weight based on the total weight of the nanoliposome.

Fatty acid to be used in nano-liposome preparation of the present invention is a higher fatty acid, preferably a C ₁₂ - ₂₂ as a saturated or unsaturated fatty acid of the alkyl chain, e.g., lauric acid, myristic acid, palmitic acid, stearic acid, oleic acid and linoleic acid . The amount thereof may be 0.05 to 3.0% by weight, preferably 0.1 to 1.0% by weight, based on the total weight of the nanoliposome.

The water used in the preparation of the nanoliposome of the present invention is generally deionized distilled water, and the amount thereof may be 5.0-40 wt% based on the total weight of the nanoliposome.

The preparation of nanoliposomes can be accomplished through a variety of methods known in the art, but most preferably is made by applying a mixture comprising the ingredients to a high pressure homogenizer. The preparation of a nanoliposome by a high pressure homogenizer can be carried out under various conditions (for example, pressure, number of times, etc.) depending on a desired particle size, and preferably at a high pressure homogenizer So that the nanoliposome can be produced.

The cosmetic composition according to the present invention may contain the bamboo leaf fermentation product in an amount of 0.1-20.0 wt% based on the weight of the nanoliposome filler, and more preferably 1.0-10.0 wt% in order to stabilize the formulation.

In one embodiment of the present invention, the bamboo leaf fermented product may be contained in the cosmetic composition at a concentration of 0.1 μg / ml to 10000 μg / ml.

In addition, the composition of the present invention may be a feed composition or feed additive capable of imparting antioxidant functionality.

The feed composition according to the present invention may contain the bamboo leaf fermentation product in an amount of 0.1 to 50% by weight, more preferably 1 to 20% by weight, based on the weight of the composition, but is not limited thereto. On the other hand, when used as a feed additive, bamboo leaf fermented product can be used in a feed composition.

Hereinafter, the present invention will be described in more detail with reference to Examples. These examples are for further illustrating the present invention, and the scope of the present invention is not limited to these examples.

< Example  1>

God of leaves Fermentation product  Produce

In this experiment, Bacillus subtilis, which is mainly used for the fermentation of food, was used to ferment Korean leaves.

The strains used for fermentation were Bacillus subtilis (KCCM 11315), which was purchased from KCCM (Seoul, Korea). TSB medium (tryptic soy broth) was used for the microbial culture and cultured at 37 ° C with shaking at 150 rpm. The bamboo leaves were dried by natural drying of Sasa coreana Nakai leaves collected in Damyang - eup, Damyang - gun, Jeollanam - do Province in early August 2016. Bacillus subtilis (KCCM 11315), which was cultured for 8 hours in a large glass bottle, was inoculated with OD600 nm 0.50 ± 0.02 using sterile 0.9% NaCl, Lt; RTI ID = 0.0 &gt; 37 C &lt; / RTI &gt; incubator.

As a result, as shown in Fig. 1, when the ginseng leaves were fermented with a strain of Bacillus subtilis (KCCM 11315), the leaves which were clearly distinguished from the rough surface were softened by the softening after the fermentation, , And it became more brown as the fermentation proceeded. Especially, phenolic compounds (polyphenols) were improved in extraction efficiency through fermentation process. In addition, various components were changed.

God leaves for the fermentation process leaves rough god for the present invention was briefly called 'SC-F (S. coreana Nakai fermented)', without passing through the process of fermentation is used as a control 'SC-N (S. coreana Nakai non-fermented '.

Each of them was naturally dried, pulverized and stored, and some of them were extracted with 80% ethanol and concentrated.

< Example  2>

General compositional changes of soybean fermentation

In this experiment, the contents of general components such as sugars, minerals,

<2-1> Analysis of composition

Crushed SC-N and SC-F were mixed with 72% sulfuric acid and 1:10 (w / v) for compositional analysis, respectively, and then extracted in a shaking incubator at 30 ° C for 2 hours. After distilled water was added to the extract, acid hydrolysis was carried out at 121 ° C for 1 hour to pre-treat the sample. For the analysis of free sugar, the crushed SC-N and SC-F were mixed with distilled water at 1:50 (w / v), respectively, and then the sample was pre-treated at 70 ° C for 5 hours. The reaction mixture was filtered through a 0.45 μm membrane filter. Carbohydrate column (250 × 4 mm, CarboPac PA-1, Dionex Co., USA) was used for the analysis. 18 mM sodium hydroxide was used as the mobile phase, and the constituents and free sugars of the extract were analyzed using a flow rate of 1.0 mL / min, a column temperature of 25 ° C, an injection volume of 25 μL, and a detector using PAD.

The detailed results are shown in Table 1 below.

The constitutional and free sugar content (mg / g) of SC-N and SC-F Party Rams North Arabinose Galactose Glucose Xylose Fructose gun Free sugar SC-N N. D. 0.15 + - 0.01 1.66 + 0.03 15.60 ± 0.89 0.26 ± 0.05 10.45 + 1.05 28.12 ± 2.03 SC-F N. D. 0.13 + 0.03 0.06 ± 0.01 N. D. 1.21 ± 0.10 N. D. 1.4 ± 0.14 Per constitution SC-N 1.44 ± 0.04 32.13 + - 0.52 18.27 ± 0.27 283.39 ± 1.99 96.86 +/- 2.56 N. D. 432.09 ± 5.38 SC-F 1.90 + - 0.12 33.76 ± 0.93 16.85 + - 0.12 291.17 ± 3.07 114.74 + - 3.15 N. D. 458.42 + - 7.39

N. D .: Not detected

As shown in Table 1, the glucose content of SC-N and SC-F was the highest at 283.39 ± 1.99 mg / g and 291.17 ± 3.07 mg / g, respectively, and that of xylose, arabinose, galactose, It was confirmed that the sugar content was high in order of osseous. The total content of constituent sugars was 432.09 ± 5.38 mg / g for SC-N and 458.42 ± 7.39 mg / g for SC-F, indicating that the constituent sugars increased after fermentation compared to before fermentation. As a result of free sugar analysis, SC-N had high contents in the order of glucose, fructose, galactose, xylose and arabinose, and only free arabinose, galactose and xylose were detected in free sugars of SC-F. The total free sugar content of SC-N was 28.12 ± 2.03 mg / g and SC-F was 1.4 ± 0.14 mg / g, which was lower than that before fermentation.

In the previous study, it was reported that the total sugar content after fermentation of both mugwort and mugwort was decreased when fermented by using Bacillus subtilis in both mugwort and mugwort (Song, HN (2013)). Rapid pattern analysis of their volatile flavor components via SAW based electronic nose sensor in the GC system. J Food Preserv 20 (4): 554-563.) In another study, (Reis, BAD, A. Kosiska-Cagnazzo, R. Schmitt, and W. Andlauer (2014) Fermentation of Saccharomyces cerevisiae strain has been reported to reduce total sugar content, including glucose and fructose, of Plant Material-Effect on Sugar Content and Stability of Bioactive Compounds. PJFNS 64 (4): 235-241.). The results of these previous studies show different results from the experimental results of the present invention. The difference is judged by the difference between the kinds of leaves and inoculated microorganisms and the fermentation conditions, and the reason why the composition of sugar is changed is that the microorganism , It is very likely that sugar was used as a growth promoting factor.

<2-2> Mineral Analysis

After adding 0.5 mL of SC-N and SC-F to 7 mL of 65% nitric acid (HNO ₃ ) and 1 mL of 30% hydrogen peroxide, the microwave digestion system (ETHOS TC, Milestone, Sorisole, Italy) ). &Lt; / RTI &gt; After cooling the acid dissociation reaction solution, it was cooled to 50 mL with 1% nitric acid, filtered through a 0.45 μm membrane filter, and analyzed for minerals. The minerals were analyzed using ICP-OES (OPTIMA 7300 DV, Perkin Elmer, USA), and the Rf power was 1400 W, the gas flow rate was 15 L / min, the nebulizer gas flow was 0.65 L / min, (auxillary gas flow) at 0.2 L / min.

Mineral analysis was performed on 9 kinds of Fe, Zn, P, Na, K, Ca, Mg, Cu and Mn. The detailed results are shown in Table 2 below.

Mineral content (μg / mL) of SC-N and SC-F Minerals SC-N SC-F Fe 90.1 ± 2.8 78.3 + - 4.7 Zn 13.3 ± 2.0 17.2 ± 2.3 P 3782.3 ± 26.0 2541.0 ± 58.1 Na 23.2 ± 0.9 203.3 ± 8.9 K 11680.7 ± 386.2 12332.7 ± 316.7 Ca 3435.3 ± 34.1 4136.3 ± 32.0 Mg 1534.3 ± 38.1 1403.0 + - 115.2 Cu 4.1 ± 0.1 3.5 ± 0.6 Mn 240.6 ± 7.2 272.3 + - 42.4 Total 20804.1 + - 364.6 20987.5 ± 345.1

As shown in Table 2, the total mineral content of SC-N 20804.1 ± 364.6 μg / mL and SC-F 20987.5 ± 345.1 μg / mL increased the total mineral content after fermentation, but there was no significant difference. After fermentation, the contents of minerals were highest at 12332.7 ± 316.7 μg / mL in K, followed by Ca, P and Mg, and Ca and P and Mg decreased. Na increased about 8.76 times (180.1 μg / mL), which is thought to be due to the effect of 0.95% NaCl (saline water) used to maintain the osmotic pressure during the inoculation of Bacillus subtilis strains used for fermentation.

The changes of mineral contents were considered to be caused by the fermentation of Bacillus subtilis strains used for fermentation, which facilitated the diffusion of minerals as the leaves softened.

<2-3> Amino acid analysis

The constituent amino acids were extracted by hydrolyzing the crushed SC-N and SC-F with hydrochloric acid at 110 ° C for 24 hours. Free amino acids were extracted with 80% ethanol. The extracts were prepared at 10,000 ppm using methanol and then transferred to a new vial by completely transferring 30 μL of the extract to 20 mL of a derivatization solution (MeOH: H2O: TEA: PITC = 7: 1: 1: 1) And then derivatized at room temperature for 30 minutes. The derivatized samples were dried again and then dissolved in 200 μL of 140 mM sodium acetate (NaHAc), 0.1% triethylamine (TEA) and 6% acetonitrile (CH ₃ CN), filtered through a 0.45 μm membrane filter Were used for analysis. HPLC was performed with Hewlett Packard 1100 Series (Agilent Co., USA) and Nove-Pak (300 × 3.9 mm, 4.0 μm, C18, Waters Co., USA) was used. For the mobile phase, 140 mM NaHAc, 0.1% TEA, 6% CH ₃ CN (solution A) and 60% CH ₃ CN (solution B) 13%, 12%, 13-15.2 minutes 20%, 15.2-22.5 minutes 46%, 22.5-22.72 minutes 100%, 22.72-25.7 minutes 100%, 25.7-26 minutes 0%, 26-30 minutes 0%. Amino acids were analyzed at a flow rate of 1.0 mL / min, a column temperature of 46 ° C, an injection volume of 10 μL, and a UV detector at a wavelength of 254 nm.

Analysis of the constituent amino acids and free amino acids was performed on a total of 20 amino acids, and the detailed results are shown in Tables 3 and 4 below.

Total amino acid content of SC-N and SC-F (mg / 100 g) SC-N SC-F Cysteine & cystine 1133.84 + - 6.81 3648.61 ± 5.12 Asparagine & Aspartic acid 6013.94 + - 11.62 7260.32 +/- 14.53 Glutamine & Glutamic acid 7727.87 ± 17.57 8396.07 ± 22.83 Serine 3513.73 + - 8.10 3695.28 + - 4.58 Glycine 1089.43 + - 10.05 4385.75 8.01 Histidine N. D. N. D. Arginine 835.56 ± 1.90 2586.01 + - 5.94 Threonine 927.69 ± 2.06 2903.74 + - 8.71 Alanine 6417.36 + - 23.04 4316.11 + - 12.52 Proline 3125.70 ± 9.33 4465.65 +/- 7.50 Tyrosine 684.59 + - 4.13 1543.56 ± 3.87 Balin 3344.51 + - 17.30 3958.34 + 8.41 Methineonine N. D. 50.46 + - 0.46 Isoleucine 2331.91 + 15.39 2967.57 ± 3.42 Leucine 1334.09 ± 5.24 4941.49 ± 11.00 Phenylalanine 13019.00 ± 23.52 11841.05 ± 51.85 Tryptophan 3703.35 ± 16.66 4754.50 ± 12.49 Lysine 3268.60 ± 14.93 2217.90 + - 6.00 Total essential amino acids 27922.49 + - 66.63 33635.05 ± 79.65 Total amino acid 58464.51 ± 109.12 73932.41 + 137.13

N. D .: Not detected

Free amino acid content of SC-N and SC-F (mg / 100 g) SC-N SC-F Aspartic acid 2368.37 ± 18.89 357.67 ± 2.08 Glutamic acid 450.17 + - 5.03 1447.17 + - 8.25 Asparagine 1885.33 + - 4.51 N. D. Serine 2915.67 ± 17.21 N. D. Glutamine 3194.67 ± 18.23 N. D. Glycine 356.01 + - 4.39 N. D. Histidine N. D. N. D. Arginine 448.70 + - 3.6 N. D. Threonine 713.17 ± 3.25 686.33 + - 8.5 Alanine 5986.67 + 27.23 150.17 + 4.19 Proline 2722.33 + - 22.5 N. D. Tyrosine 2058.33 ± 17.56 271.33 + - 8.08 Balin 2650.33 + -9.7 N. D. Methionine 139.33 + - 2.52 N. D. Cysteine 1926.83 + - 7.29 N. D. Isoleucine 1445.33 + - 8.08 N. D. Leucine 1082.33 + - 10.5 N. D. Phenylalanine 1755.33 + - 6.43 1915.17 ± 13.42 Tryptophan 476.67 + - 3.06 N. D. Lysine 207.10 ± 1.65 N. D. Total essential amino acids 8469.60 ± 24.59 2601.50 ± 8.79 Total free amino acids 32782.67 ± 92.49 4827.83 + - 19.76

N. D .: Not detected

As shown in Table 3 and Table 4, the total content of constitutive amino acids of SC-N was 58464.51 ± 109.12 mg / 100 g, and the total content of essential amino acids was 27922.49 ± 66.63 mg / 100 g. In contrast, the total content of constitutive amino acids of SC-F was 73881.94 ± 137.59 mg / 100 g, and the total content of essential amino acids was 33584.59 ± 80.11 mg / 100 g. The content of phenylalanine was highest in both fermentation and fermentation, followed by glutamine, glutamic acid, asparagine and aspartic acid. The total free amino acid content of SC-N was 32782.67 ± 92.49 mg / 100 g, and the total content of essential amino acids was 8469.60 ± 24.59 mg / 100 g. In contrast, the total free amino acid content of SC-F was 4827.83 ± 19.76 mg / 100 g and the total content of essential amino acids was 2601.50 ± 8.79 mg / 100 g. Amino acid contents of free amino acids decreased by more than 85% after fermentation, and glutamic acid and phenylalanine were increased compared to before fermentation. Bacillus sp. And Aspergillus sp., Which are used for the fermentation of food, are known to decompose proteins contained in foods into small molecule peptides and amino acids by generating strong proteases (Kim, JI, MJ Kang, and TW Kwon (2003) Antidiabetic effect of soybean and chongkukjang. Korea Soybean Digest 20: 44- 52.). Although there are many reports that free amino acids were increased after fermentation, the results of this experiment showed conflicting results. This is because the inoculated Bacillus subtilis (KCCM 11315) secretes protease and consumes amino acid as nitrogen source.

< Example  3>

Changes of polyphenols and catechins due to the fermentation of Shin leaves

In this experiment, total polyphenol content, total flavonoid content and total catechin content of Shin -

<3-1> Total polyphenol and total flavonoid content

The representative antioxidant components of plants are vitamins A, C, and E along with flavonoids, carotenoids, and phenolic compounds. They have excellent radical scavenging effects and have physiological activities such as anticancer and antioxidant activity. Polyphenols, the most abundant, are phenolic compounds with two or more aromatic rings, such as flavanoides and tannins, and are known to be produced by plants to protect themselves from extreme environments as they grow. These phenolic compounds are antioxidants and can protect against various diseases such as regulation of reactive oxygen species (ROS) present in the human body, prevention of oxidative damage, inhibition of inflammatory activity, and exist in many leaves and fruits of plants.

In this experiment, we investigated the antioxidative activity of Bacillus subtilis fermented by fermentation of Bacillus subtilis, and investigated the changes of polyphenol and flavonoid contents.

Total polyphenol content (TPC) was measured using the modified Folin-Denis method (Kahkonen, MP, AI Hopia, HJ Vuorela, JP Rauha, K. Pihlaja, TS Kujala, and M. Heinonen plant extracts containing phenolic compounds. J. Agric. Food Chem. 47: 39543962.). The concentrated extract was used at a concentration of 1,000 ppm by using methanol. 1,000 ppm of extract, 0.2 N Folin and Ciocalteu's phenol reagent, and 2% sodium carbonate were mixed in the same amount and reacted at room temperature for 30 minutes. The absorbance of the solution reacted at 750 nm wavelength was measured using a UV-VIS spectrophotometer. TPC is expressed as the standard concentration of gallic acid (GAE * mg / g).

The total flavonoid content (TFC) was measured by changing the method of Chang (2002) (Chang, CC, MH Yang, HM Wen, and JC Chern (2002) Estimation of Total Flavonoid Content in Propolis by Two Complementary Colorimetric Methods. Drug Anal 10 (3): 178-182.). The concentrated extract was used at a concentration of 1,000 ppm by using methanol. 0.5 mL extract, 1000 mL of methanol, 0.1 mL of 10% aluminum chloride, 0.1 mL of 1 M potassium acetate, and 2.8 mL of distilled water were mixed and reacted at room temperature for 1 hour. The absorbance of the solution reacted at a wavelength of 415 nm using a UV-VIS spectrophotometer was measured. The TFC is expressed as a standard substance quercetin reference concentration QE * mg / g.

As a result, total polyphenol content (TPC) of SC-N and SC-F was 25.51 ± 1.04 GAE * mg / g and 35.34 ± 0.91 GAE * mg / g, respectively, (TFC) were 69.64 ± 0.26 QE * mg / g and 80.57 ± 0.22 QE * mg / g, respectively, indicating that the TPC and TFC of the larval leaves were increased after the fermentation of Bacillus subtilis strains. These changes before and after fermentation are thought to be due to hydrolytic enzymes produced by Bacillus subtilis strains.

<3-2> Total catechin content

A typical catechin among flavonoids is contained in a large amount in tea leaves, and together with polyphenols, it acts effectively to remove reactive oxygen species and can act as an antioxidant. In this experiment, changes in the content of carboxychine as a compound capable of exhibiting antioxidant activity were investigated.

Total catechin content (TCC) was analyzed by HPLC (SPD-20A, SIMADZU Co., Japan). (-) - epicatechin, (-) - epigallocatechin-3-gallate, (-) - gallocatechin-3-gallate were analyzed by methanol- gallate, (-) - epicatechin-3-gallate and (-) - catechin-3-gallate were purchased from Cerilliant Co., USA. Shim-pack GIS-ODS (C18, 4.6 x 250 mm, 5.0 μm, SIMADZU Co., Japan) was used as the column. Solution A was dissolved in methanol (solution B) and methanol (solution B) was used as a mobile phase. Solution B was stored at 0% for 0-1 min, 63% for 1-45 min, 45-50 min 0% and 50-60 minutes 0%, respectively. Catechin was analyzed at a flow rate of 1.0 mL / min, a column temperature of 30 ° C, an injection volume of 30 μL, and a UV detector wavelength of 280 nm. TCC is the total catechin content of the catechin mixture.

The detailed results are shown in Table 5 below.

Catechin content of SC-N and SC-F (mg / L) SC-N SC-F (+) - Catechin 9.59 + - 0.46 19.88 ± 0.91 (-) - Epigallocatechin-3-gallate 0.39 + - 0.12 0.47 + 0.07 (-) - Epicatechin 34.10 ± 0.87 20.72 ± 1.04 (-) - Gallocatechin-3-gallate 3.62 ± 0.21 5.51 + - 0.23 (-) - Epicatechin-3-gallate 10.43 + - 0.56 3.42 ± 0.20 (-) - Catechin-3-gallate 11.50 ± 0.58 8.23 ± 0.69 Total 69.64 + - 0.94 58.23 + - 0.76

(+) - catechin, (-) - epicatechin and (-) - gallocatechin-3-gallate of SC-N and SC-F were increased and the content of (-) - epigallocatechin-3 (-) - epicatechin-3-gallate and (-) - catechin-3-gallate were decreased. On the other hand, total catechin content (TCC) was found to be SC-F 58.23 ± 0.76 mg / L after fermentation at SC-N 69.64 ± 0.94 mg / L before fermentation and decreased after fermentation of Bacillus subtilis strain.

The reason for this decrease in catechin content after fermentation is that during fermentation catechins produce dimeric compounds such as theaflavin and thearubigins by polyphenol oxidase (Y. Zhang, Y. Zhang, Y. Zhang, Q. Peng, and Z. Lin (2016) Study of the dynamic changes in the non-volatile chemical constituents of black tea during fermentation processing by a non-targeted metabolomics approach. Food Res Int 79: 106-113.). The increased functionality of the aglycone-type polyphenols, which can easily work in the metabolism of the human body, can be expected to have higher functionality.

< Example  4>

Analysis of Antioxidant Activity of Shin-Ida Leaf Fermentation

Since changes in the total polyphenol content of plants are related to antioxidation, it is essential to confirm the antioxidant effect of bamboo leaves on changes in total polyphenol content after fermentation. Therefore, the elimination of ABTS radicals was confirmed.

The ABTS radical scavenging activity was measured by changing the method of Jeong et al. (2009) with 2,2'-azino-bis (3-ethylbenzthiazoline-6-sulfonic acid (ABTS, Sigma, USA) In Vitro Antioxidative Activities and Phenolic Composition of Hot Water Extract from Different Parts of Cudrania tricuspidata. J. Choi, J. Kim, J. Kwak, H. Heo, KH Shim, BR Cho, YI Bae and JS Choi The ABTS stock solution was diluted with PBS (pH 7.4) after mixing the same amount of 7 mM ABTS solution and 2.45 mM potassium persulfate solution. The extracts were diluted to a concentration of 50-500 ppm by using methanol and 200 μL of the extract prepared by concentration and 1,000 μL of ABTS stock solution were mixed with each other to obtain an absorbance value of 0.80 ± 0.02 at a wavelength of 730 nm After reaction in a dark room for 15 minutes, the absorbance at 730 nm was measured using a UV-VIS spectrophotometer (SCINCO, Seoul, Korea) The IC ₅₀ of the extracts showed a cleavage ability for the ABTS radicals and was shown to correspond to the removal of 50% of the radicals.

As a result, as shown in FIG. 3, the DPPH radical and the ABTS radical scavenging ability of SC-N and SC-F were confirmed, and it was confirmed that the radical scavenging ability increases with increasing concentration of the extract. That is, the IC ₅₀ of the ABTS radical scavenging ability of SC-N and SC-F was increased from 301.24 ± 8.96 μg / mL before fermentation to 187.26 ± 4.78 μg / mL after fermentation and increased after the fermentation of Bacillus subtilis strain. The IC ₅₀ value of the two radical scavenging activities of gallic acid used as a positive control was 51.77 ± 1.28 μg / mL. The increase of total antioxidant activity after the fermentation of the leaves of this ginseng was considered to be due to the increase of total polyphenol contents.

The present invention has been described with reference to the preferred embodiments. It will be understood by those skilled in the art that various changes in form and details may be made therein without departing from the spirit and scope of the invention as defined by the appended claims. Therefore, the disclosed embodiments should be considered in an illustrative rather than a restrictive sense. The scope of the present invention is defined by the appended claims rather than by the foregoing description, and all differences within the scope of equivalents thereof should be construed as being included in the present invention.

Claims (5)

As a cosmetic composition having antioxidative activity,
The composition comprises Sasa coreana Nakai leaf fermented product as an active ingredient;
The ginseng larval leaf fermented product is prepared by inoculating the dried ginseng with Bacillus subtilis to the larval leaf and fermenting at 37 DEG C for 15 days;
The ginseng larval leaf fermented product had a total polyphenol content of 35.34 ± 0.91 GAE * mg / g and a total flavonoid content of 80.57 ± 0.22 QE * mg / g;
(-) - epigallocatechin-3-galactate content was 0.47 ± 0.07 mg / L, while the (-) - epicatechin content of the freshly fermented broth was 19.88 ± 0.91 mg / The content of (-) - Galocatechin-3-galactate was 5.51 ± 0.23 mg / L and the content of (-) - epicatechin-3-galate was 3.42 ± 0.20 mg / L , And the (-) - catechin-3-galactate content is 8.23 ± 0.69 mg / L. delete delete delete delete

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